Method of using (2-imidazolin-2-ylamino) quinoxalines in treating ocular neural injury

ABSTRACT

The present invention provides a method of providing neuroprotection to a mammal comprising administering to said mammal suffering from or at risk of suffering a noxious action on its nerve cells an effective amount of a compound of formula I to inhibit or prevent nerve cell injury or death  
                 
 
     wherein the 2-imidazolin-2-ylamino group is in either the 5- or 6-position of the quinoxaline nucleus; x, y and z are in any of the remaining 5-, 6-, 7- or 8-positions and are selected from hydrogen, halogen, lower alkyl, lower alkoxy or trifluoromethyl; and R is an optional substituent in either the 2- or 3-position of the quinoxaline nucleus and may be hydrogen, lower alkyl or lower alkoxy, or pharmaceutically acceptable salts thereof and mixtures thereof. Such noxious action may result from ischemia, e.g. spinal ischemia.

CROSS REFERENCE TO RELATED APPLICATIONS

[0001] This patent application is a continuation of U.S. patentapplication Ser. No. 09/655,579 filed Sep. 6, 2000 which is acontinuation of U.S. patent application Ser. No. 09/225,036 filed Jan.4, 1999 now U.S. Patent No. 6,194,415 B1 issued Feb. 27, 2001, which isa continuation-in-part of U.S. patent application Ser. No. 08/496,262filed Jun. 28, 1995 now U.S. Pat. No. 5,856,329 issued Jan. 5, 1999.

FIELD OF THE INVENTION

[0002] The present invention relates to methods for the protection ofnerve cells, including the retina, optic nerve and spinal cord ofmammals from noxious provocations including damage from compressive ormechanical effects or trauma or stress factors, including but notlimited to impaired blood flow to the nerves, and with respect to theretina and optic nerve, glaucoma, retinitis pigmentosa, and age-relatedmacular degeneration.

BACKGROUND OF THE INVENTION

[0003] Glaucoma is a disease of the eye characterized at least initiallyby increased intraocular pressure. On the basis of its etiology,glaucoma has been classified as primary or secondary. Primary glaucomais an independent syndrome in adults may be classified as either chronicopen-angle or chronic angle-closure. Primary open angle glaucoma is themost commonly occurring form of glaucoma where there is no otherattributable underlying cause. Angle-closure glaucoma usually afflictsthose persons having “shallow” angles in the anterior chamber andresults from the sides (or angels) of the chamber coming together andblocking aqueous outflow through the trabecular meshwork. Secondaryglaucoma, as the name suggests, results from pre-existing oculardiseases such as uveitis, intraocular tumor or enlarged cataract.

[0004] The underlying causes of primary glaucoma are not yet well known.Increased intraocular pressure can be a result of obstruction of aqueoushumor outflow. In chronic open-angle glaucoma, the anterior chamber andits anatomic structures appear normal, but drainage of the aqueous humoris impeded. In acute and chronic angle-closure glaucoma, the anteriorchamber is shallow, the filtration angle is narrowed and the iris mayobstruct the trabecular meshwork at the entrance to the canal ofSchlemm. Dilation of the pupil may push the root of the iris forwardagainst the angle or may produce pupillary block and thus precipitate anacute attack of elevated intraocular pressure. Eyes with narrow anteriorchamber angles are predisposed to acute angle-closure glaucoma attacksof varying degrees of severity.

[0005] Secondary glaucoma is caused by any interference with the flow ofaqueous humor from the posterior chamber into the anterior chamber and,subsequently, into the canal of Schlemm. Inflammatory disease of theanterior segment may prevent aqueous escape by causing completeposterior synechia in iris bombe, and may plug the drainage channel withexudates. Other common causes are intraocular tumors, enlargedcataracts, ventral retinal vein occlusion, trauma to the eye, operativeprocedures and intraocular hemorrhage.

[0006] Considering all types together, glaucoma occurs in about 2% ofall persons over the age of 40 and may be asymptomatic for years beforeprogressing to rapid loss of vision. It is not clear whetherglaucomatous nerve damage is the end result of one pathological processor whether there are several mechanisms by which the final disease ismanifest.

[0007] There is growing evidence that more than one pathomechanism maybe involved early in the glaucomatous process. See for example: Ruben,S. T., Hitchings, et al., Eye 8 (5) pp 516-20 (1994). Among those riskfactors are elevated intraocular pressure, family history of glaucoma,age and the vertical cup-to-disk ratio of the internal structures in theposterior chamber of the eye. One study found that in hypertensive eyeswithout visual field loss, the most important factors in predicting thelikelihood of glaucoma-induced loss were the cup-to-disk ratio and age.Johnson, C. A., Brandt, J. D., et al., Arch. Ophthalmol. 113(1) pp.70-76 (1995). These studies implicitly assume that there are persons whohave elevated intraocular pressure (ocular hypertension) without nervedamage to the optic disk or the retina. See also: Pfeiffer N., Bach, M.Ger. J. Ophthalmol. 1(1) pp. 35-40 (1992). Glaucomatous field damage isalso known to occur in the eyes of individuals with normotensiveintraocular pressure. One theory is that the size of the optic diskdetermines the susceptibility of the nerve head to glaucomatous visualfield damage at statistically normal intraocular pressure. Burk, R. O.,Rohrschneider, K., Noack, H., et al. Graefes Arch. Clin. Exp.Ophthalmol. 230 (6) pp. 552-60 (1992). Another explains visual fielddamage at normotensive pressure as occurring by a different, as yetunidentified, pathologic mechanism. Trick, G. L., Doc. Ophthalmol. 85(2) pp. 125-33 (1993). Regardless of the theory, glaucomatous visualfield damage at statistically normal intraocular pressure is aclinically recognized condition.

[0008] Elevated intraocular pressure, while being generally acknowledgedas a risk factor for the possible onset of glaucoma, is not a necessarycondition for glaucomatous field damage. Nerve cell damage can occurwith or without elevated intraocular pressure and nerve cell damage doesnot necessarily occur in individuals who experience elevated intraocularpressure. Two studies have suggested that increased choroidal perfusion(circulation) may help to prevent glaucomatous optic nerve damage inpatients with ocular hypertension. Schmidt, K.G., von Ruckmann, A., etal., Ophthalmologica, 212 (1) pp. 5-10 (1998) and Kerr J.; Nelson P.;O'Brien C., Am, J Ophthalmol., 126 (1) pp. 42-51 (1998). Thus, modernlyit appears that glaucoma is characterized as a complex syndrome thatmanifests itself as optic nerve damage with or without elevatedintraocular pressure. It further appears that each symptom, eitherelevated intraocular pressure or glaucomatous damage to nerve cells, canoccur independently of the other. The present invention provides methodsto protect retinal ganglion cells and the optic nerve that are damagedor lost despite a therapeutic lowering of intraocular pressure to withinnormal levels; to protect such cells from damage in the case ofso-called normotensive glaucoma; and to protect such cells inglaucomatous eyes that do not respond adequately to treatment modalitiesintended to lower intraocular pressure.

[0009] In cases where surgery is not indicated, topicalbeta-adrenoceptor antagonists have been the drugs of choice for treatingglaucoma. However, alpha adrenergic agonists have more recently beenapproved for use in the treatment of elevated intraocular pressure andare probably becoming mainstays in the treatment of the disease. Amongthis class of drugs are various quinoxaline derivatives having alpha₂agonist activity which were originally suggested as therapeutic agentsby Danielewiez, et al. in U.S. Pat. Nos. 3,890,319 and 4,029,792. Thesepatents disclose compounds as regulators of the cardiovascular systemwhich have the following formula:

[0010] where the 2-imidazolin-2-ylamino group may be in any of the 5-,6-, 7- or 8-position of the quinoxaline nucleus; x, y and z may be inany of the remaining 5-, 6-, 7- or 8-positions and may be selected fromhydrogen, halogen, lower alkyl, lower alkoxy or trifluoromethyl; and Ris an optional substituent in either the 2- or 3-position of thequinoxaline nucleus and may be hydrogen, lower alkyl or lower alkoxy.The presently useful compounds may be prepared in accordance with theprocedures outlined by Danielewiez, et al. The contents of both U.S.Pat. Nos. 3,890,319 and 4,029,792 are hereby incorporated by referencein their entirety.

[0011] In “Ocular Effects of a Relatively Selective Alpha-2 Agonist(UK-14,304-18) in Cats, Rabbits and Monkeys” [J. A. Burke, et al.,Current Eye Rsrch., 5, (9), pp. 665-676 (1986)] the quinoxalinederivative shown below and having the generic name brimonidine was shownto be effective in reducing intraocular pressure in rabbits, cats andmonkeys. Compounds in this study were administered topically to thecorneas of the study animals.

[0012] It has long been known that one of the sequelae of glaucoma isdamage to the optic nerve head. The optic nerve head or optic disk iswhere, along with the retinal vasculature, the axons of the retinalganglion cell (RGC) bodies that are distributed along the upper layer ofthe retina converge and are bundled together to transmit signals to thelateral geniculate nucleus. (See diagram of FIG. 6.) Damage to the opticnerve head, clinically referred to as cupping, is observable as areas ofdepression in the nerve fiber of the optic disk. Cupping is the resultof death of optic nerve fibers and alterations in the lamina cribosa, anextracellular matrix that provides structural support. Loss ofperipheral vision is a consequence of RGC demise and usually goesundetected until more advanced stages of the disease wherein up to fiftypercent of the retinal ganglion cells may already be damaged or lost.Left untreated glaucoma can progress from dimming of vision or loss ofacuity to total blindness.

[0013] Unfortunately despite long-term lowering intraocular pressure tostatistically normal levels by administration of drugs or by surgery tofacilitate outflow of the aqueous humor, damage to the nerves inglaucomatous conditions still persists in a significant number ofpatients. This apparent contradiction is addressed by Cioffi and VanBuskirk [Surv. of Ophthalmol., 38, Suppl. p. S107-16, discussionS116-17, May 1994] in the article, “Microvasculature of the AnteriorOptic Nerve” . They state:

[0014]  The traditional definition of glaucoma as a disorder ofincreased intraocular pressure (IOP) oversimplifies the clinicalsituation. Some glaucoma patients never have higher than normal IOP andothers continue to develop optic nerve damage despite maximal loweringof IOP.

[0015] The fact that the nerve damage associated with glaucoma mayprogress even after significant reduction of intraocular pressure hasled many to suggest that pressure-independent causes contribute in manycases. See for example: Schulzer M. et al., “Biostatistical evidence fortwo distinct chronic open-angle glaucoma populations” Br. J. Ophthal. pp74916-74200 (1990); Lamping K A, et al., “Long-term evaluation ofinitial filtration surgery” Ophthalmology 93 (1) pp. 91-101 (1986);Migdal, 1994; Spaeth G L “Proper outcome measurements regardingglaucoma: the inadequacy of using intraocular pressure alone.” Eur. J.Ophthal. 6 (2) pp 101-105 (1996). These causes have been suggested toinclude: (1) induction of apoptosis (programmed cell death) of retinalganglion cells which is a genetically controlled process wherebyunneeded or damaged cells die without eliciting an inflammatory response(see for example: Quigley H A, et al. Invest. Ophth. Vis. Sci., 36pp.774-786 (1995) “Retinal Ganglion Cell Death in Experimental Glaucomaand after Axotomy Occurs by Apoptosis”) and (2) further neuronaldegeneration affecting cells (which were not injured by the primaryinsult) after death or injury of incipiently injured nerve cells. Thedamage to nerve cells secondary to the primary injury result fromoveraccumulation of excitatory neurotransmitters released and othernoxious environmental conditions created by the death and degenerationof neighboring RGCs. More minor contributors or less understoodcomponents in glaucomatous optic neuropathy are: genetic determinantscontributing to irregularities in the metabolism of the extracellularmatrix and hence susceptibility of the RGCs to damage; vascularcompromise which promotes ischemia whether or not related to elevatedIOP; and metabolic disorders. Another advantage of the present inventionis that it provides a more direct and broader level of protection tonerves because the compounds of the present invention afford protectionat the locus of neural damage from both primary and secondary causes.

[0016] Retinitis pigmentosa is the term for a group of inheriteddiseases that affect the retina, the delicate nerve tissue composed ofseveral cell layers that line the inside of the back of the eye andcontain photoreceptor cells. These diseases are characterized by agradual breakdown and degeneration of the photoreceptor cells, theso-called rods and cones, which results in a progressive loss of vision.It is estimated that retinitis pigmentosa affects 100,000 individuals inthe United States. The rods are concentrated outside the center of theretina, known as the macula, and are required for peripheral vision andfor night vision. The cones are concentrated in the macula and areresponsible for central and color vision. Together, rods and cones arethe cells responsible for converting light into electrical impulses thattransfer messages to the retinal ganglion cells which in turn transmitthe impulses through the lateral geniculate nucleus into that area ofthe brain where sight is perceived. RP therefore affects a differentretinal cell type of those affected by glaucoma. Most common in alltypes of retinitis pigmentosa is the gradual breakdown and degenerationof the rods and cones. Depending on which type of cell is predominantlyaffected, the symptoms vary, and include night blindness, lostperipheral vision (also referred to as tunnel vision), and loss of theability to discriminate color before peripheral vision is diminished.

[0017] Symptoms of retinitis pigmentosa are most often recognized inadolescents and young adults, with progression of the disease usuallycontinuing throughout the individual's life. The rate of progression anddegree of visual loss are variable. As yet, there is no known cure forretinitis pigmentosa.

[0018] While not a cure, certain doses of vitamin A have been found toslightly slow the progression of retinitis pigmentosa in someindividuals. Researchers have found some of the genes that causeretinitis pigmentosa. It is now possible, in some families with X-linkedretinitis pigmentosa or autosomal dominant retinitis pigmentosa, toperform a test on genetic material from blood and other cells todetermine if members of an affected family have one of several retinitispigmentosa genes, and therefore to begin therapy before the damagingeffects of the disease become manifest. It is an object of the presentinvention to protect the photoreceptor cells, the rods and cones by thecompounds and methods described herein, particularly in regard to thestudies of protection of the photoreceptor cells to light induceddamaged by neuroprotective compounds.

[0019] Age-related macular degeneration (ARMD) is degenerative conditionof the macula or central retina. It is the most common cause of visionloss in the Western world in the over 50 age group. It most commonlyaffects those of northern European descent and is uncommon inAfrican-Americans and Hispanics. Its prevalence increases with age andaffects 15% of the population by age 55 and over 30% are affected by age75. Macular degeneration can cause loss of central vision and makereading or driving impossible, but unlike glaucoma, macular degenerationdoes not cause complete blindness since peripheral vision is notaffected. Macular degeneration is usually obvious during ophthalmologicexamination.

[0020] Macular degeneration is classified as either dry(non-neovascular) or wet (neovascular). In its exudative, or “wet,”form, a layer of the retina becomes elevated with fluid, causing retinaldetachment and wavy vision distortions. Abnormal blood vessels may alsogrow into, or under, the retina, creating a neovascular membrane thatcan leak, further obscuring vision. In advanced cases, scar tissueforms, causing irreversible scotomas, or blind spots. Dry maculardegeneration, although more common, typically results in less severe,more gradual loss of vision as one or more layers of the retinadegenerates and atrophies. Yellow deposits, called “drusen,” or clumpsof pigment may appear.

[0021] In both forms, the area of the retina affected is the macula(3)—the most sensitive area of the retina. For this reason, people withmacular degeneration lose central vision and the ability to see finedetail, while their peripheral vision remains unchanged.

[0022] In the case of age related macular degeneration, treatments havebeen proposed and studied but have found limited success in clinicalapplication. Laser photocoagulation is effective in sealing leaking orbleeding vessels. Unfortunately, it usually does not restore lost visionbut only slows or prevents further loss. Conventional laser treatmentfor exudative macular degeneration generally is effective for a limitedamount of time because the abnormal blood vessels tend to grow back.

[0023] A newer, investigational approach, photodynamic therapy, hasshown some promising results in the treatment of wet (neovascular) ARMD.An injection of a photosensitive dye is given systemically to a patient,which is taken up only in abnormal tissues such as the abnormal vesselspresent in wet ARMD “A “cold” laser is directed into the eye whichactivates the dye taken up in the cell walls of the abnormal vessels,thus forming oxidative compounds that lead to clot formation in theneovascular tissues. Fluid leakage is thus halted and as the remainingfluid is reabsorbed, vision improves. Unfortunately, the body alsoabsorbs the clot in 4-12 weeks, so the procedure must be repeated, and,additionally, the laser treatment can cause photic damage to the retina.Another aspect of the present invention is that the compounds of theinvention may be administered to protect the retina from damage by thelaser light used as a part of this ARMD therapy.

[0024] An invasive surgical technique also has been developed that usesspecialized forceps to enter into the eye and pull out the neovascularmembrane. Unfortunately the neovascularization often grows back.

[0025] The cells that nurture the retina, the cells of the retinalpigment epithelium, as well as photoreceptor tissues, have beenharvested from human fetaltissues grown in the laboratory and thentransplanted. In studies of rats with inherited retinal disease, humanfetal retinal pigment epithelium was surgically introduced into the eyeswhere it functioned normally and restored vision. Unfortunately,transplants in human studies, while initially successful, have failedwithin three months owing to rejection.

[0026] Thus it is evident that there is an unmet need for agents thathave neuroprotective effects that can stop or retard the progressivedamage resulting from one or more noxious provocations to nerve cells.

SUMMARY OF THE INVENTION

[0027] A new method of protecting the nerve cells of the eye and thespine of a mammal from noxious provocations has been discovered. Thismethod comprises administering to the mammal either systemically,topically, intrathecally, epidurally or by intrabulbar injection aneffective amount of one or more of certain aryl-imino-2-imidazolidines(as defined herein), salts thereof and mixtures thereof.

[0028] For treatment of glaucomatous; retin in humans suffering fromthat condition, the active compounds (or mixtures or salts thereof) areadministered in accordance with the present invention to the eye admixedwith an ophthalmically acceptable carrier. Any suitable, e.g.,conventional, ophthalmically acceptable carrier may be employed. Acarrier is ophthalmically acceptable if it has substantially no longterm or permanent detrimental effect on the eye to which it isadministered. Examples of ophthalmically acceptable carriers includewater (distilled or deionized water) saline and other aqueous media. Inaccordance with the invention, the active compounds are preferablysoluble in the carrier which is employed for their administration, sothat the active compounds are administered to the eye in the form of asolution. Alternatively, a suspension of the active compound orcompounds (or salts thereof) in a suitable carrier may also be employed.

[0029] In accordance with the invention the active compounds (ormixtures or salts thereof) are administered in an ophthalmicallyacceptable carrier in sufficient concentration so as to deliver aneffective amount of the active compound or compounds to the eye.Preferably, the ophthalmic, therapeutic solutions contain one or more ofthe active compounds in a concentration range of approximately 0.0001%to approximately 10% (weight by volume) and more preferablyapproximately 0.0005% to approximately 0.5% (weight by volume).

[0030] Any method of administering drugs directly to a mammalian eye maybe employed to administer, in accordance with the present invention, theactive compound or compounds to the eye to be treated. By the term“administering directly” is meant to exclude those general systemic drugadministration modes, e.g., injection directly into the patient's bloodvessels, oral administration and the like, which result in the compoundor compounds being systemically available. The primary effect on themammal resulting from the direct administering of the active compound orcompounds to the mammal's eye is preferably a reduction in intraocularpressure. More preferably, the active useful compound or compounds areapplied topically to the eye or are injected directly into the eye.Particularly useful results are obtained when the compound or compoundsare applied topically to the eye in an ophthalmic solution (oculardrops).

[0031] Topical ophthalmic preparations, for example ocular drops, gelsor creams, are preferred because of ease of application, ease of dosedelivery, and fewer systemic side effects, such as cardiovascularhypotension. An exemplary topical ophthalmic formulation is shown belowin Table 1. The abbreviation q.s. means a quantity sufficient to effectthe result or to make volume. TABLE I Ingredient Amount (% W/V) ActiveCompound in accordance about 0.0001 to about 1 with the invention,Preservative 0-0.10 Vehicle 0-40 Tonicity Adjustor 1-10 Buffer 0.01-10PH Adjustor g.s pH 4.5-7.5 Antioxidant as needed Purified Water asneeded to make 100%

[0032] Various preservatives may be used in the ophthalmic preparationdescribed in Table I above. Preferred preservatives include, but are notlimited to, benzalkonium chloride, chlorobutanol, thimerosal,phenylmercuric acetate, and phenylmercuric nitrate. Likewise, variouspreferred vehicles may be used in such ophthalmic preparation. Thesevehicles include, but are not limited to, polyvinyl alcohol, povidone,hydroxypropyl methyl cellulose, poloxamers, carboxymethyl cellulose,hydroxyethyl cellulose, and purified water.

[0033] Tonicity adjustors may be added as needed or convenient. Theyinclude, but are not limited to, salts, particularly sodium chloride,potassium chloride, mannitol, and glycerin, or any other suitableophthalmically acceptable tonicity adjustor.

[0034] Various buffers and means for adjusting pH may be used so long asthe resulting preparation is ophthalmically acceptable. Accordingly,buffers include but are not limited to, acetate buffers, citratebuffers, phosphate buffers, and borate buffers. Acids or bases may beused to adjust the pH these formulations as needed.

[0035] In a similar vein, ophthalmically acceptable antioxidantsinclude, but are not limited to, sodium metabisulfite, sodiumthiosulfate, acetylcysteine, butylated hydroxyanisole, and butylatedhydroxytoluene. The ophthalmic solution (ocular drops) may beadministered to the mammalian eye as often as necessary to obtain thedesired concentration intravitreally that affords neuroprotection. Foracute neuroprotective effect such as photoprotection in the abovedescribed laser treatment for ARMD, the protective agent would beadministered in advance of the treatment to provide optimal protectionduring the laser procedure. For chronic treatments such as in protectionof the retinal ganglion cells against damage from the neuropathiceffects of for example glaucoma or dry ARMD, the drug would beadministered as frequently as necessary to maintain desired intravitrealconcentration or range of concentrations at all times. In other words,the ophthalmic solution (or other formulation) which contains theα₂-adrenergic agent as the active ingredient, is administered to themammalian eye as often as necessary to maintain the beneficialneuroprotective effect of the active ingredient in the eye. Thoseskilled in the art will recognize that the frequency of administrationdepends on the precise nature of the active ingredient and itsconcentration in the ophthalmic formulation. Within these guidelines itis contemplated that the ophthalmic formulation of the present inventionwill be administered to the mammalian eye approximately once or twicedaily.

[0036] Treatment of ARMD using the compounds of the present inventiontakes a different therapeutic approach than the treatment modalitiesdiscussed above which concentrate on treating the vascularization.

[0037] Treatment with (α-adrenergic agonists protects the retinal cellsof the macula from damage caused by the noxious provocations of thedegenerative process or from damage of laser light used in thetreatment. These noxious occurrences include, but are not limited to:laser light damage from the laser procedure with or withoutphotoreactive dye, and atrophy associated with the dry form of ARMD.Thus the α₂-adrenergic agonists of the present invention may beadministered alone or in conjunction with any of the foregoing describedtherapies.

[0038] This new method is particularly effective when administered as aprophylactic treatment, i.e. before damage to the nerve has taken place,or before long-term progression of the disease state, such as glaucoma,retinitis pigmentosa or ARMD, has taken place. Without wishing to beheld to a particular theory regarding the role that the compounds of thepresent invention play in neuroprotection, applicants hypothesize thatthe compounds and methods described cause an upregulation of bFGF (aneuronal cell survival factor) expression via α₂ stimulation and thatthis endogenous release may provide neuroprotection by signaling thecells to survive notwithstanding the apoptotic (programmed cell death)signals the cells receive from the noxious provocation. With increasedendogenous concentrations of the bFGF in response to the α₂ agonism, thebalance of cell survival and cell death signals may be shifted towardsthe promotion of cell survival. Further, it has been found that certainfactors of the bcl-2 family are also produced as measured by theincreased expression of mRNA encoding their production; these factorslabeled bcl-2 and bcl-x_(L), also suppress the apoptotic program. Thesefactors can counterbalance presence or induction of bcl-2 apoptosisfactors such as bad and bax which may be produced as a result of noxiousprovocations to the nerve cells. Thus it is further contemplated thatthe compounds of the present invention which provide cell survivalsignals to the nerve can advantageously be used in combination withcompounds that inhibit cell death. Such cell death inhibiting compoundsinclude NMDA antagonists especially memantine, which block excitotoxiceffects of excess glutamate nitric oxide synthetase inhibitors;free-radical scavengers and calcium channel blockers.

DETAILED DESCRIPTION OF THE INVENTION

[0039] The drawings will first be briefly described.

DRAWINGS

[0040]FIG. 1 is a bar graph showing the percentage of cells killed bytreatment with glutamate plotted by number of days since glutamatetreatment. A control which was not treated with glutamate has beenincluded to determine cell death which occurred without any suchtreatment. Also shown are measurements taken after treatment with bothAGN191103 and glutamate, and treatment with MK-801 and glutamate.N4K-801 is a well known neuroprotective agent in the art that acts as anNMDA antagonist. The NMDA receptor binds, among other neurotransmitters,glutamate. The numbers beneath the bars for glutamate;AGN191103+glutamate; and MK-801+glutamate show the concentrations ofglutamate and drug used in each case. At day 8, AGN 191103 and MK-801show comparable effects in protecting cells from glutamate inducedneurotoxicity. Experimental procedures followed in generating the datafor this figure are detailed in Example 1.

[0041]FIG. 2 shows plots of compound action potentials (CAP) measuredfor optic nerve fibers: in the left-hand frame, measured at 2 weeks postinjury (i.e. after nerve crush) for optic nerve treated with AGN 191103(the upper line) and for an untreated nerve used as a control (lowerline); and in the right-hand frame a comparison CAP of non-injured opticnerve. The scales of the plots are given for each of the frames. Thepost-injury abscissa scale is 25× the scale of the non-injured plot.(Units: millivolts and milliseconds). The value of the compound actionpotential is calculated as the integral of the area under each curve.The irregularity of the curve feature of the dispersion of the compoundresponse; some nerve cells conduct more rapidly than others and soamplitude of the measured voltage varies with time.

[0042]FIG. 3 is a bar graph showing the maximal CAP amplitude inmicrovolts (μV) for cells injured by a optic nerve crush in rats andtreated with: 1) vehicle alone; 2) clonidine and 3) AGN 191103. Each ofthe drugs was tested at four different concentrations (administered as amultiple of body weight for the test subject) and is represented by abar on the chart. Clonidine was chosen as a benchmark α₂ agonistcompound with very well defined pharmacology to compare against the testcompound AGN 191103. While clonidine did show some neuroprotectiveactivity over vehicle alone, it showed about half the maximal CAPresponse of AGN 191013.

[0043]FIG. 4 is a graphic plot of the Visual Evoked Potential Responseand shows the electrical potential activity evoked at the surface of thevisual cortex (comparable to an electroencephalogram) as a result ofvisual (light) stimulus. The test is performed in live rats and is ameasure of the integrity of the whole visual system from the retinathrough the optic nerve into the lateral geniculate nucleus andultimately to the visual cortex located in the back of the brain. Theleft-hand frame shows the response without nerve crush injury and theright-hand frame shows the responses measured at 2 weeks post-injury forrats treated with AGN 191103 above (labeled positive) and control ratsbelow (labeled negative) prior to nerve crush. The scale in μV vs.milliseconds for both plots is shown below the ordinate axis.

[0044]FIG. 5 is a bar graph showing the results of a study to determinethe topical efficacy of brimonidine in neuroprotection. The rat acuteretinal ischemia model was used to provide the noxious action.Brimonidine was applied topically one hour before ischemia insult, 10 μlto one eye, 10 μl saline to the other eye. The ERG (electroretinogram)was measured one week after the insult. The bar graph shows the percentof ERG signal recovery as a function of dosage. The results show thatbrimonidine provides topical neuroprotection in a dose dependent manner.

[0045]FIG. 6 is a schematic drawing of a cut away section of the eyewhich shows the anterior and posterior chambers. The former being filledwith aqueous humor and separated from the posterior around the lens ofthe eye. The posterior segment is filled with vitreous humor (6) aclear, viscous liquid that maintains the shape of the eye. At the backof the eye are the retina (4), optic disk (3), and the optic nerve (5).Underlying the retina is the retinal pigment epithelium and choroid (2)which is responsible for maintenance and support of the retinal nervecells. Further the diagrammatic view of the retina shows the layers ofnerve cells and associated helper cells that make up the retina. Thefirst layer of cells contacted by light incoming through the lens is theretinal ganglion cells (7), then at the base of the retina toward thechoroid are the photoreceptor cells (8) which are comprised of the rods(9) and cones (10).

[0046] For a discussion and bibliography regarding the nerve crush modeland its significance in evaluating nerve damage and recovery see:Functional Recovery and Morphological Changes after Injury to the OpticNerve, Sabel, B. A. and Aschoff, A., Neuropsychobiology, 28, pp. 62-65(1993).

[0047] Injury to the mammalian optic nerve, as in any other parts of themammalian central nervous system (CNS), leads to axonal degenerationfollowed by a loss of cell bodies, with failure of axonal regrowth fromthe surviving neurons. Initially, degeneration of the injured nerve isprobably attributable to direct neuronal damage. However, the associatedphysiological and biochemical events occurring in the nerve immediatelyafter injury are probably responsible for the subsequent progressivedegeneration, not only of the directly injured axons, but also of thosethat escaped the primary damage. These secondary effects largelydetermine the long-term functional outcome.

[0048] The immediate injury-induced response strongly influences thesubsequent degenerative response. Treatment that reduces or attenuatesthe immediate response is therefore likely to achieve optimal preventionor delay of the secondary degenerative processes. For monitoring of theimmediate response, it is obviously preferable to employ a noninvasivetechnique. An adaptation of the nicotinamide adenine dinucleotide (NADH)monitoring technique to enable measurement of the earliestpost-traumatic events has proved to be a valuable non-invasive approach.Use of the technique allows the immediate effect of the injury to beevaluated in real time and on-line before and after a well-controlledcrush injury is inflicted on the adult rat optic nerve in vivo. In thisexperimental paradigm, measurement of the metabolic activity of theinjured optic nerves represent the activity of both injured axons andtheir associated non neuronal cells, and thus evaluate the potentialability to cope with injurious stresses. The model is also useful formonitoring the activity of various agents that may overcome or mitigatenerve cell damage or death from such stresses.

[0049] The earliest injury-induced response is a decrease in the energystate of the nerve, under conditions where ischemic events can becompletely ruled out. The reduction in the energy state may be relatedto: 1) postinjury elevation in free fatty acid levels, which mayinterfere with mitochondrial function and result in uncoupling ofelectron transport; and 2) a marked rise in intracellular free Ca²⁺. Itis known that axonal injury is generally followed by an increase inextracellular potassium ions, which stimulate the uptake of Ca²⁺ viaeither voltage sensitive channels (L, T or N type) or receptor-operatedCa²⁺ channels. A marked rise in intracellular free Ca²⁺ can accelerateprocesses that are inimical to cell survival, including those involvingCa²-dependent enzymes, mainly lipases, proteases and endonucleases, thatmay cause mitochondrial damage and lead eventually to cell death. Thecell, in order to overcome these events, needs more energy to activelyrestore ionic homeostasis. The combination of increased energy demandsand decreased energy conservation resulting from mitochondrialdysfunction at the site of injury may be the major reason for thesubsequent irreversible nerve damage and nerve degeneration followinginjury. Early measurement of metabolic activity could therefore indicatethe fate of the axon, its associated glial cells and its non-neuronalcell bodies. It follows from the above that restoration of themitochondrial activity may be critical in preventing the degenerativeprocess occurring in the nerve after injury.

[0050] Since the injury inflicted on the nerve in the nerve crush modelis a well-controlled, calibrated and reproducible lesion, it is possibleto correlate early post-traumatic metabolic deficits and possiblemitigation of these by drug or other treatments with long-termmorphological and physiological effects.

[0051] From the foregoing figures and discussion it is apparent thatneuroprotection is conferred on nerve cells against bothglutamate-induced toxicity and physical insult in the nerve crush model.

[0052] It has now been discovered that neuroprotection is conferred uponocular nerve cells by administration of a drug of formula I to thespinal neurons or retina and optic nerve of a mammal within a periodprior to, or following an insult to the nerve cells but prior to celldeath,

[0053] wherein the 2-imidazolin-2-ylamino group may be in either the 5-or 6-position of the quinoxaline nucleus; x, y and z may be in any ofthe remaining 5-, 6-, 7- or 8-positions and are selected from hydrogen,halogen, lower alkyl, lower alkoxy or trifluoromethyl; and R is anoptional substituent in either the 2- or 3-position of the quinoxalinenucleus and may be hydrogen, lower alkyl or lower alkoxy.

Definitions

[0054] The compound identified as AGN 191103 has the chemical structureas shown. It is also known by the chemical nomenclature6-methyl-(2-imidazolin-2-ylamino) quinoxaline.

[0055] The neuroprotective agent identified as MK-801 is also known bythe name dizocilpine and has the following chemical structure:

[0056] It is additionally identified and described in the 11th editionof the Merck Index at monograph number 3392.

[0057] The terms noxious actions or noxious provocations are defined asan occurrence which is harmful or destructive to a nerve cell. It is notlimited to events extrinsic to the mammal being treated but includesdisease states and physiological occurrences or events, such as, forexample, stroke or heart attack, that are harmful or destructive to thenerve cell via a chain of events. Non-limiting examples of noxiousactions include: compressive or mechanical effects or trauma or stressfactors, such as glutamate neurotoxicity, impaired blood flow to thenerves (ischemia) and with respect to the retina or optic nerves,retinitis pigmentosa and age-related macular degeneration and glaucoma.

Human Dosage and Administration

[0058] The methods of this invention are useful in treating any mammal,including humans.

[0059] According to this invention, mammals are treated withpharmaceutically effective amount of a neuroprotective agent for aperiod of time and at a time such that noxious provocations to the opticnerve and retina do not kill or permanently damage the nerve cells.Protective agents may be administered orally, topically to the eye or byany other appropriate means of delivery described below or known in theart.

[0060] In accordance with this invention, pharmaceutically effectiveamounts of a protective agent can be administered alone to treat nerveinjury or to prevent nerve cell death. Alternatively a protective agentmay be administered sequentially or concurrently with an antiglaucomadrug, e.g. beta-blocker, an alpha₂ agonist, a muscarinic agent such aspilocarpine, a carbonic anhydrase inhibitor (CAI), or another druguseful in maintaining intraocular pressure (IOP) at normal levels or inlowering elevated IOP. The most effective mode of administration anddosage regimen of protective agent will depend on the type of disease tobe treated, the severity and course of that disease, previous therapy,the patient's health status, and response to the drug and the judgmentof the treating physician. Generally, the neuroprotective agent shouldbe administered in a dose to achieve a serum or intravitrealconcentration of 0.01 nM to 500 nM. Preferably the neuroprotective agentis administered prior to injury to the nerve, but can be administeredafter injury has occurred with lessened effect.

[0061] Conventional modes of administration and standard dosage regimensof protective agents, e.g. MK-801, can be used. Optimal dosages forco-administration of a drug, e.g. an IOP-lowering drug, with aneuroprotective agent can be determined using methods known in the art.Dosages of neuroprotective agents may be adjusted to the individualpatient based on the dosage of the drug with which the agent iscoadministered and the response of the patient to the treatment regimen.The protective agent may be administered to the patient at one time orover a series of treatments.

[0062] The agent may be administered locally, e.g. intravitreally byintrabulbar injection for ocular neuroprotection, or by intrathecal orepidural administration for spinal protection. Many of the agents of theinvention can be administered systemically, e.g., orally, orintravenously, or by intramuscular injection. Additionally, agents forprotection of the retina and optic nerve that are capable of passingthrough the cornea, and achieving sufficient concentration in thevitreous humor, such as AGN 191103 and brimonidine, may also beadministered topically to the eye.

[0063] The composition used in these therapies may also be in a varietyof forms. These include, for example, solid, semi-solid, and liquiddosage forms, such as tablets, pills, powders, preserved ornon-preserved liquid solution or suspension, liposomes, suppositories,injectable and infusible solutions. The compositions also preferablyinclude conventional pharmaceutically acceptable carriers which areknown to those of skill in the art.

[0064] The following non-limiting examples describe assays andmeasurements used in 1) determining protection of nerve cells fromglutamate induced toxicity and 2) methods of determining neuralprotection conferred by neuroprotective agents in a nerve crush model ofmechanical injury.

EXAMPLE 1 Experimental Procedure for Measuring Neural Protection in aModel of Glutamate Induced Excitotoxic Effects on Nerve Cells

[0065] Low-density rat hippocampal neuronal cultures were prepared bythe procedure of Goslin and Banker. Coverslips were cleaned andsterilized in porcelain racks in such a way that they did not stick toone another (Cohen cover glass staining racks, Thomas Scientific).Coverslips (13 mm) were placed in staining racks, rinsed in distilledwater (four rinses, 1 min. each) to remove dust and transferred toconcentrated HN0 ₃ for 36 hours. Coverslips were rinsed in distilledwater (four changes over 3 hours) and sterilized with dry heat(overnight at 225° C.). The coverslips were transferred to 24-welldishes, one coverslip per well. To support the coverslips above the gliaduring coculturing, paraffin dots were placed on dishes, and UVirradiation (30 min.) was applied before the coverslips were introduced.One mg/mL of poly-L-lysine hydrobromide (PLL) (Sigma) (MW 30,000-70,000)was dissolved in borate buffer (0.1 M, pH 8.5), filtered, sterilized andused to cover each coverslip overnight. The PLL was removed, coverslipswere rinsed in distilled water (two washes, 2 hrs. each), plating medium[Eagle's MEM with Earle's salts containing extra glucose (600 mg/L) and10% horse serum] was added and the dishes were stored in an incubator.Astroglial cultures were prepared from the brains of neonatal rats by amethod similar to that described by Levinson and McCarthy, except thatthey were plated at a lower density so that they contained predominantlytype 1 astroglia. 10⁵ cells were plated in each well. Glial cultureswere fed with plating medium twice a week and were used after reachingconfluence, about 2 weeks after plating. One day before use, the platingmedium was removed, neuronal maintenance medium (MEM containing N2supplements) was added, and incubation continued. 3×104 of viable rathippocampal nerves (E18 embryos) were plated on the PLL-treatedcoverslips kept in plating medium. After 3-4 hrs, when most of theneurons were attached, the coverslips were transferred to the dishescontaining the glial cell in maintenance medium in such a way that theneuronal side was facing the glia, which support neuronal survival anddevelopment. To reduce glial proliferation, cytosine arabinoside(1-b-D-arabinofuranosylcytosine)(Calbiochem)(5×10 M final concentration)was added to the cultures 2 days after plating. At day 6 in culture,cells were treated with 1 mM glutamate or with glutamate together witheither AGN-191103−0.1 nM (MW=200) or MK-801−10 nM (2-3 coverslips wereused to each treatment).

[0066] After 24 hrs. of incubation, cells were stained with trypan blue.Live and dead neurons were counted from randomly selected culture fields(5 fields from each coverslip). Percentage of dead cells was calculated.

EXAMPLE 2 Procedure for Nerve Crush Injury and Measurements of CompoundAction Potentials (CAP) Subsequent to Injury

[0067] Part A.

Metabolic Measurements

[0068] Animal utilization was according to the ARVO Resolution on theuse of animals in research. Male Sprague-Dawley (SPD) rats weighing300-400 g were anesthetized with sodium pentobarbitone(intraperitoneally, 35 mg/kg). A cannula was introduced into the tracheafor artificial ventilation when required. With the animal's head held inplace by a head holder, a lateral canthotomy was performed under abinocular operating microscope and the conjunctiva was incised lateralto the cornea. After separation of the retractor bulbi muscles, theoptic nerve was identified and a length of 03.5 mm was exposed near theeyeball by blunt dissection. The dura was left intact and care was takennot to injure the nerve. The first part of a light guide holder (seebelow) was inserted under the optic nerve and the nerve was gently easedinto the light guide canal. The second part was then fixed in place insuch a way that the light guide was located on the surface of the opticnerve 1 mm from the site at which the injury was to be administered.

Surface Fluorometry-reflectometry

[0069] Monitoring of the intramitonchodrial NADH redox state was basedon fluorescence of NADH at 366 nm, resulting in the emission of bluelight with a peak intensity at 450 run, which is unlike its oxidizedform, NAD+, which lacks this fluorescence. The source of the 366 nmexcitation is a 100-W air-cooled mercury lamp equipped with a strong366-nm filter (Corning 5860 (7-37) plus 9782 (4-96)). A flexibleY-shaped bundle of optic fibers (light guide) is used to transmit thelight to and from the optic nerve, thus making in vivo measurementstechnically feasible. Excitation light is transmitted through the bundleof excitation fibers to the nerve. The light emitted from the nerve,after being transmitted through a second bundle of fibers, is split in aratio of 90:10 for measurement of the fluorescent light (90%) at 450 nmand the reflected light (10%) at 366 nm by two photomultipliersconnected to a one-channel direct current fluorometer-reflectometer. Inorder to minimize variations among animals, standard signal calibrationprocedures were applied at the start of the recordings. Changes in thefluorescence and reflectance signals during the experiment arecalculated relative to the calibrated signals. This type of calibration,although not absolute, has nevertheless been found to yield reliable andreproducible results from various animals and among differentlaboratories.

[0070] Changes in reflected light were correlated with changes in tissueabsorption caused by hemodynamic effects and movements of the opticnerve secondary to alteration in arterial blood pressure and nervevolume. The fluorescence measurements are found to be adequatelycorrected for NADH redox state measurements by subtraction of thereflected light (366 run) from the fluorescent light (1:1 ratio) toobtain the corrected fluorescence signal.

Metabolic Measurements

[0071] Animals which were still anesthetized were allowed to recover for30 min. from the surgical procedures described above and were thenexposed to anoxic and hyperoxic conditions. An anoxic state was achievedby having the rat breathe in an atmosphere of 100% nitrogen for 2 min.,after which it was returned to air. Whenever animals did not returnspontaneously to normal breathing, they were ventilated by blowing twiceinto the trachea. A hyperoxic state was induced by having the animalbreathe 100% oxygen for 6-10 min. In order to evaluate the metabolicactivity of the optic nerve, the relative changes in reflected andfluorescent light intensities in response to anoxia and to hyperoxiawere measured before and after crush injury.

Experimental Protocol for Metabolic Measurements

[0072] Using calibrated cross-action forceps, a well-calibrated moderatecrush injury was inflicted to the nerve between the eye and the lightguide holder at a pressure corresponding to 120 g for 30 sec.

[0073] Part B.

Physiological Measurements

[0074] Experimental setup for recording compound action potential (CAP):Prior to removal of optic nerves for electrophysiological measurement,the rats were deeply anesthetized with 70 mg/kg pentobarbitone. The skinwas removed from the skull and the optic nerves were detached from theeyeballs. Subtotal decapitation was performed and the skull was openedwith a rongeur. The cerebrum was displaced laterally, exposing theintracranial portion of the optic nerve. Dissection at the level of thechiasm enabled removal of the whole length of the nerve, which wastransferred to vials containing fresh, cold Krebs solution, consistingof: NaCl (125 mM), KCl (5 mM), KH₂PO₄ (1.2 mM), NaHCO₃ (26 mM), MgSO₄(0.6 mM), CaCl₂ (24 mM), D-glucose (11 mM), aerated with 95% O₂ and 5%CO₂. The nerves were kept in this solution, in which electrical activityremained stable for at least 3-4 h. After 1 h of recovery, nerves wereimmersed in Krebs solution at 37° C. Electrophysiological recording wereobtained from the nerve distal to the crush lesion, since the nerveswere too small to allow measurement on both sides of the crush. Thenerve ends were then connected to two suction Ag-AgCl electrodesimmersed in the bathing solution. The stimulating pulse was appliedthrough the electrode at the proximal end and the action potential wasrecorded by the distal electrode.

[0075] A Grass SD9 stimulator was used for electrical stimulation (2 V,50 μs). The signal was transmitted to a Medelec PA63 preamplifier andthence to a Medelec MS7 electromyograph and AA7T amplifier. Thesolution, stimulator and amplifier had a common ground. The maximumamplitude of eight averaged CAPs was recorded and photographed with aPolaroid camera. The left nerves (uninjured) were used to measure thereference values of normal nerves and to calibrate the crush forceps.

Recording of Visual Evoked Potential (VEP) Response

[0076] Injured drug-treated rats were examined in 2 weeks after theinjury for assessment of their functional recovery. In this set ofexperiments, the pattern of filed potentials in response to lightstimulation was recorded from the primary visual cortex. The potentialevoked by the light originates in the retina and is propagated along thesurviving axons, to reach their final target, the visual cortex. Onlythose axons that survived the primary and secondary degenerativeprocesses are capable of conducting an action potential. A comparativeanalysis of the pattern of field potentials in treated and untreatedanimals will reveal the effect of the treatment on axonal survival.

[0077] Anesthetized rats (Rumpon, Ketalar) were placed in a small animalsterotaxic instrument. After exposure of the skull, two holes weredrilled with a cylindrical drill bit, with the dura kept intact tominimize cortical damage. One hole, drilled above the nasal bone, wasused as a reference point. The second hole was in the area OC1 with thecoordinates Bregma #8 mm, lateral #3 mm. A gold contact pin connected toa screw was used as the electrode, which was screwed into the holes andglued by acrylic cement to the skull. The field potential was evoked bystroboscopic stimulation, with an average of 90 sweeps per minute. Theflash-evoked potential was analyzed by the use of the Lab View dataacquisition and management system. The field potentials were digitizedand stored for offline analysis.

[0078] Part C.

Measurement of Effects of Drug Tests for Neural Protective Properties

[0079] The first set of experiments involved metabolic measurements.Each drug was injected intraperitoneally at several differentconcentrations. Each drug was tested in a group of 8 animals, togetherwith 8 controls (injured animals treated with the buffer vehicle). Ineach case, metabolic measurements were obtained on-line before injury,0.5 h after injury and every hour for 4-6 h thereafter. The dataobtained were analyzed by ANOVA.

Measurement of Long Term Effects, Physiological Activities. CAPS

[0080] Immediately after injury, the drug to be tested was injected into10 animals, and 10 control animals were injected with vehicle. Two weekslater the CAPs of each nerve were recorded in vitro, using suctionelectrodes. The contralateral side was used as an internal control. Theresults indicated whether the examined drug had any potential effects onthe rescue of spared axons and/or slowing of degeneration. Positiveresults led to efforts at determining the optimal dosage for eachpromising drug.

VEP Response

[0081] Electrodes were implanted in the cortex of naive SPD rats in twoage- and sex-matched groups. Immediately after implantation, the VEPresponse was recorded from the left side while a light was flashed intothe right eye, with the left eye covered. A well-controlled crush injurywas then inflicted on the optic nerve and the drug was immediatelyadministered at the previously determined optimal dosage. Controlanimals were handled in the same way except vehicle was administeredrather than drug. The VEP response for each animal was recorded 1 day, 1week, 2 weeks and 4 weeks after operation.

[0082] In a similar vein, it has been demonstrated that nerve celldamage occurs in models of spinal ischemia, and suggested that inducedspinal hypothermia has neuroprotective effect. Marsala, M, Gulik, J.,Ishikawa, T. and Yaksh, T. L., Journal of Neuroscience Methods 74, pp97-106 (1997). The mechanism of nerve cell death following ischemia isbelieved to be by mechanisms similar to those found to be effectivelytreated by the administration of compounds of the present invention innerves such as the optic nerve and retina. Hypoxic neuronaldepolarization and glutamate toxicity are cited in the study by Marsala,Galik, et al (supra).

[0083] Recent studies have shown that adrenergic α_(2A), receptors arethe predominant α₂ subtype localized in the human spinal cord. It hasbeen suggested previously that α_(2A), agonists may be useful inanalgesia or sedation by binding to and activation of α_(2A) receptorsin the spine. See for example, Lawhead, R. G., Blaxall, H. S., andBylund, D. B. Anesthesiology, 77(5) 983-91 (1992). It is newly foundthat these agonists will impart neuroprotection to the cells of thespine. Administration of an effective amount of the compounds of theinvention for neuroprotection in the spine may be made intrathecally,epidurally or systemically, such as orally or by injection, as discussedabove under “Dosage and Administration”. Such administration to thespinal cord will protect spinal nerve cells from noxious provocationssuch as ischemia and trauma.

[0084] Animal models of spinal ischemia; methods of inducing suchischemia, methods for measuring amounts of endogenous compounds releasedduring and after ischemia using intrathecal dialysis and histologicalstudies of the nerve tissues and behavioral neurologic function afterischemic events are provided in the following papers: Marsala, M.,Malmberg, A. B., and Yaksh, T. L., J. Neuroscience Methods, 62, pp.43-53 (1995); Taira, Y.,Marsala, M., Stroke, 27 (10), pp. 1850-58(1996); Marsala, M., Vanicky, I., Yaksh, T. L., Stroke, 25 (10), pp.2038-46 (1994). Using the combination of these references an experimentis used to test the effectiveness of the compounds of the invention. Thecompounds are administered to a test animal, for example a rat, followedby induction of spinal ischemia and then reperfusion, or calibratednerve crush damage. Measurement of release of glutamate, and otherendogenous compounds is made by use of intrathecal dialysis. Also thestudy of neurologic behavior modifications, such as induction ofallodynia and loss of motor abilities is made to demonstrate theusefulness of the compounds for spinal neuroprotection against a controlgroup of animals that are not treated with neuroprotective compounds.

[0085] While this invention has been described with respect to variousspecific examples and embodiments, it is to be understood that theinvention is not limited thereby and should only be construed byinterpretation of the scope of the appended claims.

What is claimed is:
 1. A method of providing neural protection to amammal comprising administering to said mammal suffering from or at riskof suffering a noxious action on its nerve cells an effective amount ofa compound of formula I to inhibit or prevent nerve cell injury or death

 or pharmaceutically acceptable salts thereof and mixtures thereof. 2.The method of claim 1 wherein the noxious action is a result of acrushed or compressed nerve.
 3. The method of claim 1 wherein thenoxious action is a result of ischemia.
 4. The method of claim 1 whereinthe noxious action is a result of spinal ischemia.
 5. The method ofclaim 1 wherein the noxious action is glaucomatous optic neuropathy. 6.The method of claim 1 wherein the noxious action is atrophy associatedwith dry ARMD.
 7. The method of claim 1 wherein said noxious action is alaser light directed into the eye in a procedure for treatment of wetARMD.
 8. The method of claim 1 wherein the noxious action is ischemiasecondary to glaucoma.
 9. The method of claim 1 wherein the noxiousaction is photoreceptor cell damage associated with retinitispigmentosa.
 10. The method of claim 4 wherein said compound isadministered intrathecally, epidurally or systemically.
 11. The methodof claim 5 wherein said compound is administered orally or by injection.12. The method of claim 1 wherein said noxious action is a result ofglutamate induced excitotoxic effects on nerve cells.
 13. The method ofclaim 1 wherein said compound is administered in combination with anNMDA antagonist.
 14. The method of claim 8 wherein said NMDA antagonistis memantine.
 15. The method of claim 1 wherein said compound isadministered in an amount sufficient to achieve a serum concentration offrom 0.01 nM to 500 nM.
 16. The method of claim 1 wherein said compoundis administered topically.